EP2265613A1 - Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof - Google Patents

Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof

Info

Publication number
EP2265613A1
EP2265613A1 EP08873110A EP08873110A EP2265613A1 EP 2265613 A1 EP2265613 A1 EP 2265613A1 EP 08873110 A EP08873110 A EP 08873110A EP 08873110 A EP08873110 A EP 08873110A EP 2265613 A1 EP2265613 A1 EP 2265613A1
Authority
EP
European Patent Office
Prior art keywords
pyrrolo
methoxy
oxy
piperazinoquinazoline
benzodiazepin
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
EP08873110A
Other languages
German (de)
French (fr)
Other versions
EP2265613B1 (en
Inventor
Ahmed Kamal
Bandari Rajendra Prasad
Adla Malla Reddy
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Council of Scientific and Industrial Research CSIR
Original Assignee
Council of Scientific and Industrial Research CSIR
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Council of Scientific and Industrial Research CSIR filed Critical Council of Scientific and Industrial Research CSIR
Publication of EP2265613A1 publication Critical patent/EP2265613A1/en
Application granted granted Critical
Publication of EP2265613B1 publication Critical patent/EP2265613B1/en
Not-in-force legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to quinazoline linked pyrrolo[2,1-c][1,4] benzodiazepine hybrids and a process for the preparation thereof. More particularly it relates to 7-methoxy-8- ⁇ substituted(4-piperazinoquinazoline)alkyl]oxy ⁇ -(11aS)- 1 ,2,3, 11a-tetrahydro-5H-pyrrolo[2,1-c][1 ,4]benzodiazepine-5-one with aliphatic chain length variations useful as anticancer agent.
  • the structural formula of these quinazoline linked pyrrolo[2,1-c][1 , ⁇ benzodiazepines hybrids is given below.
  • Pyrrolo[2,1-c][1 , ⁇ benzodiazepine antitumour antibiotics are commonly known as anthramycin class of compounds.
  • PBDs pyrrolo[2,1-c][1 ⁇ benzodiazepines
  • antibiotics react covalently with DNA to form an N2-guanine adduct that lies within the minor groove of duplex DNA via an acid-labile aminal bond to the electrophilic imine at the N10-C11 position
  • PBD dimers have been developed that comprise of two C2-exo- methylene substituted DC-81 subunits tethered through their C-8 position via an inert propanedioxy linker (Gregson, S. J.; Howard, P. W.; Hartely, J. A.; Brooks, N. A.; Adams, L. J.; Jenkins, T. C; Kelland, L. R. and Thurston, D. E. J. Med. Chem. 2001, 44, 737).
  • a non-cross-linking mixed imine-amide PBD dimers have been synthesized that have significant DNA binding ability and potent antitumour activity (Kamal, A.; Ramesh, G.
  • Naturally occurring pyrrolo[2,1-c][1 , ⁇ benzodiazepines belong to a group of antitumour antibiotics derived from Streptomyces species. Recently, there is much impetus for the PBD systems as they can recognize and bind to specific sequence of DNA. Examples of naturally occurring PBDs include anthramycin, DC-81 , tomaymycin, sibiromycin and neothramycin.
  • the main objective of the present invention is to provide novel quinazoline linked pyrrolo[2,1-c][1 , ⁇ benzodiazepine hybrids, useful as antitumour agents.
  • Yet another object of this invention is to provide a process for the preparation of novel quinazoline linked pyrrolo[2,1-c][1 , ⁇ benzodiazepine hybrids.
  • the present invention provides a novel quinazoline linked pyrrolo-[2,1-c][1, ⁇ benzodiazepine hybrid of general formula 5
  • novel quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung (Hop-62), cervix (SiHa), breast (MCF7, Zr-75-1), colon (Colo205), prostate (DU 145, PC3) and oral (DWD, HT1080) cell lines.
  • the concentration of quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid used for in vitro activity against Colo205 for IC50 is in the range of 12 to 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against DU 145 for IC50 is in the range of 15 to 80 ⁇ m, at an exposure period of at least 48 hrs. In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 , ⁇ benzodiazepine hybrids used for in vitro activity against DWD for IC50 is in the range of 6 to 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 , ⁇ benzodiazepine hybrids used for in vitro activity against HoP62 for IC50 is in the range of 13 to 40 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against HT1080 for IC50 is in the range of 6 to 30 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against MCF7 for IC50 is in the range of 23 to 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against PC3 for IC50 is in the range of 5 to about 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against SiHa for IC50 is in the range of 19 to about 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against Zr-75-1 for IC50 is in the range of 16 to about 80 ⁇ m, at an exposure period of at least 48 hrs.
  • the present invention further provides a pharmaceutical composition
  • a pharmaceutical composition comprising quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid, its derivatives, analogues, salts or mixture thereof optionally with pharmaceutically acceptable carriers, adjuvants and additives.
  • quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid used is represented by a general formula 5,
  • R 1 , and R 2 is selected from the group of consisting of H, H or CH 3 O, CH 3 O or CH 3 O, C 2 H 5 O, in the presence of a mild inorganic base, in an aprotic organic solvent, under refluxing temperature to obtain the resultant nitro compound of formula 3,
  • the mild inorganic base used in steps (a) is selected from the group of potassium carbonate and Acetone.
  • the mild inorganic base used in steps (a) is potassium carbonate.
  • aprotic organic solvent used in step (a) is acetone and acetonitrile
  • organic solvent used in step (c) is acetonitrile and acetone
  • the alcohol used in step (b) is selected from methanol and ethanol.
  • the compounds of formula 5 obtained are represented by a group of the following compounds: 7-Methoxy-8- ⁇ [-3-(4-piperazinoquinazoline]propyl ⁇ -oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-][1 ,4]benzodiazepin-5-one (5a);
  • the quinazoline linked pyrrOlo[2,1-c][1 ⁇ benzodiazepine hybrid of formula 5a-o exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung, cervix, breast, colon, prostate and oral cell lines.
  • the present invention provides a process for preparation of pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids of formula 5
  • the compound 3b has been prepared according to the method described for the compound 3a by employing the compounds 1b (535 mg, 1 0 mmol) was added anhydrous K 2 CO 3 (690 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3b (608 mg, 90%).
  • the compound 4b has been prepared according to the method described for the compound 4a by employing compound 3b (669 mg, 1 0 mmol) to afford compound 4b (640 mg, 95%), which, due to potential stability problems, was used directly in the next step.
  • the compound 5b was prepared according to the method described for the compound 5a employing the compound 4b (640 mg, 1 0 mmol) to afford the compound 5b (435 mg, 67%).
  • the compound 4c has been prepared according to the method described for the compound 4a by employing compound 3c (683 mg, 1 0 mmol) to afford compound 4c (654 mg, 95%), which, due to potential stability problems, was used directly in the next step.
  • the compound 5c was prepared according to the method described for the compound 5a employing the compound 4c (655 mg, 1 0 mmol) to afford the compound 5c (450 mg, 68%).
  • the compound 3d has been prepared according to the method described for the compound 3a by employing the compounds 1d (563mg, 1 0 mmol) was added anhydrous K2CO 3 (690 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3d (655 mg, 89%).
  • the compound 4d has been prepared according to the method described for the compound 4a by employing compound 3d (698 mg, 1 0 mmol) to afford compound 4d (675 mg, 96%), which, due to potential stability problems, was used directly in the next step.
  • the compound 5d was prepared according to the method described for the compound 5a employing the compound 4d(668 mg, 1 0 mmol) to afford the compound 5d (465 mg, 69%).
  • the compound 3e has been prepared according to the method described for the compound 3a by employing the compounds 1e (563mg, 1 0 mmol) was added anhydrous K 2 CO 3 (712 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3e (675 mg, 90%).
  • the compound 4e has been prepared according to the method described for the compound 4a by employing compound 3e (714 mg, 1 0 mmol) to afford compound 4e (682 mg, 95%), which, due to potential stability problems, was used directly in the next step.
  • 5a, 5c and 5d was evaluated for in vitro anticancer activity against sixty human tumor cells derived from nine cancer types ⁇ leukemia, non-small-cell lung, colon, CNS, melanoma, ovarian, prostate, and breast cancer) as shown in (Table 1 and 2).
  • dose response curves for each cell line were measured at a minimum of five concentrations at 10 fold dilutions.
  • a protocol of 48 h continuous drug exposure was used and a sulforhodamine B (SRB) protein assay was used to estimate cell viability or growth.
  • SRB sulforhodamine B
  • the mean graph midpoint values of log 10 TGI and log-io LC50 as well as logTM GI50 for 5a is listed in Table 1 and 2). As demonstrated by mean graph pattern, compound 5c and 5d exhibited an interesting profile of activity and selectivity for various cell lines. The mean graph mid point of log-io TGI and log 1Q LC50 showed similar pattern to the log-to GI50 mean graph mid points.
  • each cancer type represents the average of six to eight different cancer cell lines.
  • In vitro evaluation of cytotoxic activity Among them 5a exhibits a wide spectrum of activity against sixty cell lines in nine cell panels, with GI 50 value of ⁇ 20 nM.
  • the growth of HOP-62, NCI-H23 cell lines were affected by compound 5a with GI 50 values as 11.4, 14.6 and 19.7 nM respectively.
  • the GI 50 values of compound 5a against colon cancer HCC-2988, HCT-116 and KM12 cell lines are 11.3, 11.8 and 11.7 nM respectively.
  • the Gl 50 values for compound 5a against CNS SF-295, SF-539, SNB-19 and SNB-75 cell lines are in a range of 12.6-23.9 nM.
  • Four cancer cell lines (OVCAR-4, OVCAR-5, OVCAR-8 and SK-OV-3) in the ovarian cancer cell panel were affected by compound 5a with Gl 50 values of 31.6, 15.1 , 30.5 and 79.8 nM respectively.
  • compound 5c exhibited cytotoxicity activity against renal and breast cancer panels with Gl 50 values (11.6-43.4 nM).
  • Compound 5c exhibits activity against sixty cell lines in nine cancer cell panels with GI 50 values of ⁇ 10 ⁇ M.
  • Compound 5d exhibits activity against fifty-seven cell lines in nine cancer cell panels, GI 50 values of ⁇ 10 ⁇ M. Cytotoxicity of compounds 5a, 5c and 5d in selected cancer cell lines has been illustrated in and 5d have been, illustrated in Table 1, 2 and 3.Table 2. The average GI 50 values for each cancer panel of compounds 5a, 5c, and 5d have been illustrated in Table 1, 2 and 3.
  • the mean graph mid point values of logTM TGl and log 10 LC 50 as well as logTM GI 5 0 for 5a, 5c and 5d are listed in Table-3. As demonstrated by mean graph pattern, compounds 5a, 5c and 5d exhibit an interesting profile of activity and selectivity for various cell lines.
  • the mean graph mid points of logTM TGI and logTM LC 50 have shown similar pattern to the logTM Gl 50 mean graph mid points.
  • logTM Gl 50 , logTM TGI and logTM LC 50 mean graphs midpoints (MG_MID) of In vitro cytotoxicity data for the compounds 5a, 5c and 5d against human tumor cell lines.
  • ADR Adiramycin is the control drug
  • CT-DNA duplex-form calf thymus DNA
  • Working solutions in aqueous buffer (10 mM NaH 2 PO 4 ZNa 2 HPO 4 , 1 mM Na 2 EDTA, pH 7.00+0.01) containing CT-DNA (100 ⁇ m in phosphate) and the PBD (20 ⁇ m) were prepared by addition of concentrated PBD solutions in DMSO to obtain a fixed [PBD]/[DNA] molar ratio of 1 :5.
  • the DNA-PBD solutions were incubated at 37 0 C for 0, 18, and 36 h prior to analysis.
  • the present invention provides a new pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids useful as antitumour agents.

Abstract

The present invention provides a compound of general formula 5, useful as potential antitumour agents against human cancer cell lines. The present invention further provides a process for the preparation of pyrrolo[2,1-c][1, 4]benzodiazepine hybrids of general formula (5): wherein n = 3, 4, 5, 6, 8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O.

Description

"QUINAZOLINE LINKED PYRROLO[2,1-q[1.^BENZODIAZEPINE HYBRIDS AS POTENTIAL ANTICANCER AGENTS AND PROCESS FOR THE PREPARATION THEREOF"
FIELD OF THE INVENTION
The present invention relates to quinazoline linked pyrrolo[2,1-c][1,4] benzodiazepine hybrids and a process for the preparation thereof. More particularly it relates to 7-methoxy-8-{substituted(4-piperazinoquinazoline)alkyl]oxy}-(11aS)- 1 ,2,3, 11a-tetrahydro-5H-pyrrolo[2,1-c][1 ,4]benzodiazepine-5-one with aliphatic chain length variations useful as anticancer agent. The structural formula of these quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepines hybrids is given below.
Formula 5 and where n=3-6,8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O
BACKGROUND OF THE INVENTION
Pyrrolo[2,1-c][1 , ^benzodiazepine antitumour antibiotics are commonly known as anthramycin class of compounds. In the last few years, a growing interest has been shown in the development of new pyrrolo[2,1-c][1 ^benzodiazepines (PBDs). These antibiotics react covalently with DNA to form an N2-guanine adduct that lies within the minor groove of duplex DNA via an acid-labile aminal bond to the electrophilic imine at the N10-C11 position (Kunimoto, S.; Masuda, T.; Kanbayashi, N.; Hamada, M.; Naganawa, H.; Miyamoto, M.; Takeuchi, T.; Unezawa, H. J. Antibiot, 1980, 33, 665.; Kohn, K. W. and Speous, C. L. J. MoI. Biol., 1970, 51, 551.; Hurley, L. H.; Gairpla, C. and Zmijewski, M. Biochem. Biophys. Acta., 1977, 475, 521; Kaplan, D. J. and Hurley, L. H. Biochemistry, 1981, 20, 7572). The molecules have a right-handed twist, which allows them to follow the curvature of the minor groove of B-form double-stranded DNA spanning three base pairs. A recent development has been the linking of two PBD units through their C-8 positions to give bisfunctional-alkylating agents capable of cross-linking DNA (Thurston, D. E.; Bose, D. S.; Thomson, A. S.; Howard, P. W/, Leoni, A.; Croker, S. J.; Jenkins, T. C; Neidle, S. and Hurley, L. H. J. Org. Chem. 1996, 61, 8141).
anthramycin
DC-81 dimers (n = 3-5); DSB-120 (n = 3) imine-amide PBD dimers; n = 3-5
Recently, PBD dimers have been developed that comprise of two C2-exo- methylene substituted DC-81 subunits tethered through their C-8 position via an inert propanedioxy linker (Gregson, S. J.; Howard, P. W.; Hartely, J. A.; Brooks, N. A.; Adams, L. J.; Jenkins, T. C; Kelland, L. R. and Thurston, D. E. J. Med. Chem. 2001, 44, 737). A non-cross-linking mixed imine-amide PBD dimers have been synthesized that have significant DNA binding ability and potent antitumour activity (Kamal, A.; Ramesh, G. Laxman, N.; Ramulu, P.; Srinivas, O.; Neelima, K.; Kondapi, A. K.; Srinu, V. B.; Nagarajaram, H. M. J. Med Chem. 2002, 45, 4679). Recently, some new pyrrolobenzodiazepine (PBD) hybrids have been synthesized that have significant DNA binding ability and potent antitumour activity.( Kamal, A.; Srinivas,' O.; Ramulu, P.; Ramesh, G.; Kumar, P. P. Bioorg. Med. Chem. Lett. 2003, 13, 3577).
Naturally occurring pyrrolo[2,1-c][1 , ^benzodiazepines belong to a group of antitumour antibiotics derived from Streptomyces species. Recently, there is much impetus for the PBD systems as they can recognize and bind to specific sequence of DNA. Examples of naturally occurring PBDs include anthramycin, DC-81 , tomaymycin, sibiromycin and neothramycin.
However, the clinical efficacy for these antibiotics is hindered by several limitations, such as poor water solubility, cardio toxicity, development of drug resistance and metabolic inactivation.
OBJECTIVES OF THE INVENTION
The main objective of the present invention is to provide novel quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepine hybrids, useful as antitumour agents.
Yet another object of this invention is to provide a process for the preparation of novel quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepine hybrids.
SUMMARY OF THE INVENTION
Accordingly the present invention provides a novel quinazoline linked pyrrolo-[2,1-c][1, ^benzodiazepine hybrid of general formula 5
wherein n = 3-6, 8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O1 R2= CH3O or R1= CH3O and R2=C2H5O
Formula 5 In an embodiment of the present invention the novel quinazoline linked pyrrolo [2,1-c][1 ,4]benzodiazepine hybrid as claimed in claim 1 is represented by the group of the following compounds:
7-Methoxy-8-{[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5a)
7-Methoxy-8-{[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)-1 ,2,3,11a-5/-/- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5b);
7-Methoxy~8-{[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5c);
7-Methoxy-8-{[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5d);
7-Methoxy-8-{[-8-(4-piperazinoquinazoline]octyl}-oxy-(11 aS)-1 ,2,3, 11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5e);
7-Methoxy-8-{6,7-dimethoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5f); 7-Methoxy-8-{6,7-dimethoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5g); 7-Methoxy-8-{6,7-dimethoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5h); 7-Methoxy-8-{6,7-dimethoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5i); 7-Methoxy-8-{6,7-dimethoxy[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5j);
7-Methoxy-8-{7-ethoxy,6-methoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5k); 7-Methoxy-8-{7-ethoxy,6-methoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5I); 7-Methoxy-8-{7-ethoxy,6-methoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)- 1 ,2,3, 11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5m); 7-Methoxy-8-{7-ethoxy,6-methoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5n); -Methoxy-8-{7-ethoxy,6-methoxy [-8-( 4-piperazinoquinazoline ]octyl}-oxy-(1 IaS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5o) In yet another embodiment the structural formula of the representative compounds of quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepine hybrids are:
π = 3,4,5,6,8
Formula 5a-e
n = 3,4,5,6,8
Formula 5f-j
n = 3,4,5,6,8 Formula 5k-o
In yet another embodiment the novel quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung (Hop-62), cervix (SiHa), breast (MCF7, Zr-75-1), colon (Colo205), prostate (DU 145, PC3) and oral (DWD, HT1080) cell lines.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid used for in vitro activity against Colo205 for IC50 is in the range of 12 to 80μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against DU 145 for IC50 is in the range of 15 to 80μm, at an exposure period of at least 48 hrs. In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 , ^benzodiazepine hybrids used for in vitro activity against DWD for IC50 is in the range of 6 to 80μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 , ^benzodiazepine hybrids used for in vitro activity against HoP62 for IC50 is in the range of 13 to 40 μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against HT1080 for IC50 is in the range of 6 to 30μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against MCF7 for IC50 is in the range of 23 to 80μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against PC3 for IC50 is in the range of 5 to about 80μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against SiHa for IC50 is in the range of 19 to about 80μm, at an exposure period of at least 48 hrs.
In yet another embodiment the concentration of quinazoline linked pyrrolo[2,1-c] [1 ,4]benzodiazepine hybrids used for in vitro activity against Zr-75-1 for IC50 is in the range of 16 to about 80μm, at an exposure period of at least 48 hrs.
The present invention further provides a pharmaceutical composition comprising quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid, its derivatives, analogues, salts or mixture thereof optionally with pharmaceutically acceptable carriers, adjuvants and additives.
In yet another embodiment the quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid used is represented by a general formula 5,
Formula 5 wherein n = 3, 4, 5, 6, 8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2O2H5O The present invention further provides a process for the preparation of quinazoline linked pyrroio[2,1-c][1, ^benzodiazepine hybrid of formula 5
wherein n = 3, 4, 5, 6, 8. and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O
The said process comprising the steps of: a) reacting(2S)-/V-[(n-bromoalkyloxy)-5-methoxy-2-nitrobenzoyl)]pyrrolidine- 2-carboxaldehyde diethylthioacetal of formula 1
1a-e where n = 3 to 6 and 8 with nonsubstituted and substituted-4-piperazinoquinazoline derivative selected from the compound of formula 2a-c
2a-c wherein R1, and R2 is selected from the group of consisting of H, H or CH3O, CH3O or CH3O, C2H5O, in the presence of a mild inorganic base, in an aprotic organic solvent, under refluxing temperature to obtain the resultant nitro compound of formula 3,
3a-o wherein R1 and R2 is selected from the group consisting of R1= H1 R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O
reducing the above said nitro compound of formula 3 obtained in step (a) with SnCb.2H2O in an organic solvent, under reflux temperature and isolating the corresponding amino compound of formula 4,
wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O1 R2= CH3O or R1= CH3O and R2=C2H5O c) reacting the above said amino compound of formula 4 obtained in step (b) with a deprotecting agent in a mixture of water and organic solvent in the presence of a mild inorganic basic compound by known method to obtain the desired compound of formula 5.
In yet another embodiment the mild inorganic base used in steps (a) is selected from the group of potassium carbonate and Acetone.
In yet another embodiment the mild inorganic base used in steps (a) is potassium carbonate.
In yet another embodiment the aprotic organic solvent used in step (a) is acetone and acetonitrile
In yet another embodiment the organic solvent used in step (c) is acetonitrile and acetone
In yet anther embodiment the alcohol used in step (b) is selected from methanol and ethanol.
In yet another embodiment the compounds of formula 5 obtained are represented by a group of the following compounds: 7-Methoxy-8-{[-3-(4-piperazinoquinazoline]propyl}-oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-][1 ,4]benzodiazepin-5-one (5a);
7-Methoxy-8-{[-4-( 4-piperazinoquinazoline ]butyl}-oxy-(11aS)-1 , 2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5b);
7-Methoxy-8-{[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5c);
7-Methoxy-8-{[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5d);
7-Methoxy-8-{[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5e);
7-Methoxy-8-{6,7-dimethoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)- 1 ,2,3, 11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5f); 7-Methoxy-8-{6,7-dimethoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5g); 7-Methoxy-8-{6,7-dimethoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)- 1 ,2,3,11a-5/-/-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5h); 7-Methoxy-8-{6,7-dimethoxy[-6-(4-piperazinoquinazoline]hexyI}-oxy-(11aS)- 1 ,2,3, 11 a-5H-pyrrolo[2, 1-c][1 ,4]benzodiazepin-5-one (5i); 7-Methoxy-8-{6,7-dimethoxy[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5j); 7-Methoxy-8-{7-ethoxy,6-methoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)- 1 ,2,3, 11 a-5H-pyrrolo[2,1 -c][1 ,4]benzodiazepin-5~one (5k); 7-Methoxy-8-{7-ethoxy,6-methoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5l); 7-Methoxy-8-{7-ethoxy,6-methoxy[-5-(4-piperazinoquinazoline]pentyI}-oxy-(11aS)- 1 ,2,3,11 a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5m); 7-Methoxy-8-{7-ethoxy,6-methoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5n);
7-Methoxy-8-{7-ethoxy,6-methoxy [-8-( 4-piperazinoquinazoline ]octyl}-oxy-(1 IaS)- 1 ,2,3, 11 a-5H-pyrrolo[2, 1 -C][1 ,4]benzodiazepin-5-one (5o)
In still another embodiment the quinazoline linked pyrrOlo[2,1-c][1 ^benzodiazepine hybrid of formula 5a-o exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung, cervix, breast, colon, prostate and oral cell lines.
DETAILED DESCRIPTION OF THE INVENTION
Accordingly, the present invention provides a process for preparation of pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids of formula 5
where n = 3-6, 8, and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O and which comprises reacting substituted and unsubstituted A- piperazinoquinazoline derivative of formula 2 with (2S)-Λ/-[(n-bromoalkyloxy)I5- methoxy-2-nitrobenzoyl)]pyrrolicline-2-carboxaldehyde diethylthioacetal of formula 1 in presence of CH3COCH3/K2CO3 for a period of 48 h resulting in isolation of (2S)- Λ/-{n-[substituted or un-substituted(4-piperazinoqui"':izoline]alkyl-oxy-5-methoxy-2- nitrobenzoyl}pyrrolidi-ne-2-carboxaIdehyde diethyl thioacetal of formula 3 by conventional methods. Reducing the above nitro compound of formula 3 with SnCl2.2H2θ in presence of organic solvent with reflux temperature, resulting with the formation of (2S)-Λ/-{n-[4-piperazinoquinazoline]alkyloxy]}-5-methoxy-2- aminobenzoyl}pyrrolidine-2-carboxaldehyde diethylthioacetal 4. Reacting the above said amino compound of formula 4 with known deprotecting agents in a conventional manner to give novel pyrrolo[2,1-c][1,4]benzodiazepine hybrids, of formula 5, where 'n' is as stated above.
The precursors, substituted 4-piperazinoquinazoline of formula 2(Liou, J. P.; Chang, C.W.; Song, J. S.; Yang, Y. N.; Yeh, C. F.; Tseng, H. Y.; Lo, Y. K.; Chang, Y. L.; Chang, C. M.; Hsieh, H. P.; J. Med. Chem. 2002, 45, 4513-4523). and (2SJ- Λ/-(4-hydroxy-5-methoxy-2-nitrobenzoyl)pyrrolidine-2-carboxaldehyde diethylthioacetal of formula 1 (Thurston, D. E.; Morris, S. J.; Hartley, J. A. Chem. Commun. 1996, 563-565) have been prepared by literature methods.
Some representative compounds of formula 5 for the present inventions are given below
(5a)7-Methoxy-8-{4-[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one.
(5b)7-Methoxy-8-{4-[-3-( 4-piperazinoquinazoline]butyl}-oxy-(11aS)-1 ,2,3,11a-5/-/- pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one.
(5c)7-Methoxy-8-{4-[-3-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one.
(5d)7-Methoxy-8-{4-[-3-(4-piperazinoquinazoline]hexyl}-oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one.
(5e)7-Methoxy-8-{4-[-3-(4-piperazinoquinazoline]octyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one.
The process for the preparation of new pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids is disclosed and claimed in our co-pending Indian patent application no.
603/DEL/2008.
These new analogues of pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids linked at C-8 position have shown promising DNA binding activity and efficient anticancer activity in various cell lines. The molecules synthesized are of immense biological significance with potential sequence selective DNA-binding property. This resulted in design and synthesis of new congeners as illustrated in Scheme-1 , which comprise:
1. The ether linkage at C-8 position of DC-81 intermediates with 3-(4-hydroxy- 3-methoxyphenyl)-1-(4-phenyl-3-quinolyl)-2-propen-1-one moiety.
2. Refluxing the reaction mixtures for 48 h.
3. Synthesis of C-8 linked PBD antitumour antibiotic hybrid imines.
4. Purification by column chromatography using different solvents like ethyl acetate, hexane, dichloromethane and methanol.
Reagents and conditions: (i) SOCI2, C6H6, L-prolinemethyl ester hydrochloride, THF-H2O, 2 h, rt, 85%, (n) DIBAL-H, CH2CI21I h, -78 0C, 71%, (in) EtSH, TMSCI, CH2CI2, 8 h, rt, (ιv) EtSH-BF3OEt2, CH2CI2, 12 h, rt, 75%, (v) dibromoalkanes, K2CO3, acetone, 48 h, refllux, 94-96%, (vi) compound 3, acetonitπle, 48 h, refllux, 94-96%, (VIi) SnCI2 2H2O, MeOH, 2 h, reflux, 85-87%, (vili) HgCI2-CaCO3, CH3CN-H2O (4 1), 12 h, rt, 68-71 %
The following examples are given by way of illustration and therefore should not be construed to the present limit of the scope of invention.
Example-1
(2S)-Λ/-{4-[3-[4-Piperazinoquinazoline]propyl]oxy]-5-methoxy-2-nitrobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (3a)
To a solution of compound (2S)-Λ/-[4-(3-Bromopropoxy)-5-methoxy-2- nitrobenzoyl]pyrrolidine-2-carboxaldehyde diethylthioacetal (1a) 521 mg, 1.0 mmol) in dry acetonitrile (15 ml) was added, 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) followed by anhydrous K2CO3 (690 mg, 2.0 mmol) and the reaction mixture was refluxed for 48 hrs. After the completion of the reaction as indicated by TLC, EtOAc:hexane (7:3), the reaction mixture was concentrated under reduced pressure and then extracted with ethyl acetate. This was concentrated under reduced pressure to obtain the crude product which was further purified by column chromatography on silica gel eluting with EtOAc:hexane (9:1) to obtain the pure compound 2S)-Λ/-{4-[3-[4-Piperazinoquinazoline]propyl]oxy]-5-methoxy-2- nitrobenzoyl}pyrrolidine-2-carboxal- dehyde diethylthioacetal 3a as yellow solid (560 mg, 85%).
1H NMR (CDCl3) δ 1.32-1.40 (m, 6H), 1.75-2.30 (In, 6H), 2.61-2.86 (In, 6H), 3.21- 3.30 (m, 6H), 3.80-3.86 (t, 4H / = 7.32 Hz), 3.96 (s, 3H), 4.19-4.24 (t, 2H), 4.66-4.74 (m, IH), 4.85-4.87 (d, IH, / = 3.96), 6.82 (s, IH), 7.42-7.47 (t, IH, / = 6.63 Hz), 7.70-7.75 (m, 2H), 7.86-7.91 (m, 2H); 8.71 (s, IH); MS (FAB) 655.27 [M+H].
(2S)-Λ/-{4-[3-[4-Piperazinoquinazoline]propyl]oxy]-5-methoxy-2-aminobenzoyl} pyrroIidine-2-carboxaldehyde diethylthioacetal (4a)
The compound 3a (655 mg, 1 Ommol) dissolved in methanol (20 ml) and added SnCI2.2H2O (1.125 g, 5 0 mmol) was refluxed for 2 h or until the TLC indicated that reaction was complete. The methanol was evaporated under vacuum and the aqueous layer was then adjusted to pH 8 with 10% NaHCO3 solution and then extracted with ethyl acetate (2x30 ml). The combined organic phase was dried over Na2SO4 and evaporated under vacuum to afford the amino diethyl thioacetal, 4a, which, due to potential stability problems, was used directly in the next step (612 mg, 97%).
7-Methoxy-8-{3-[4-piperazinoquinazoline]propyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one(5a)
A solution of 4a (625 mg, 1 0 mmol), HgCI2 (678 mg, 2.5 mmol) and CaCO3 (250 mg, 2.5 mmol) in acetonitrile-water (4:1) was stirred at room temperature for 12 h until the completion of the reaction as shown by TLC (EtOAc). The organic layer was evaporated in vacuum and the residue was diluted with EtOAc. To this, saturated NaHCO3 was added slowly at room temperature and the mixture was filtered through a celite bed and washed with ethyl acetate. The filtrate was evaporated in vacuum to get crude compound 5a, which was further purified by column chromatography on silica gel eluting first with ethyl acetate to remove traces of mercuric salts and then with CHCI3:methanol (9:1) (392 mg, 62%). 1H NMR (CDCl3) δ 1.25-1.42 (m, 4H), 1.64-1.88 (m, 4H), 2.02-2.20 (m, 3H), 2.28-2.39 (m, 2H), 2.63-2.78 (m, 4H), 3.80-3.91 (m, 2H), 3.96 (s, 3H), 4.18-4.24 (t, 2H, / = 6.73 Hz)), 7.42-7.56 (m, 2H), 7.67-7-79 (m, 2H), 7.87-7.94 (m, 2H), 8.74 (s, IH); MS (FAB) 500 [M+H]+
Example-2
(2S)-Λ/-{4-[4-[4-piperazinoquinazoline]butyl]oxy]-5-methoxy-2-nitrobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (3b)
The compound 3b has been prepared according to the method described for the compound 3a by employing the compounds 1b (535 mg, 1 0 mmol) was added anhydrous K2CO3 (690 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3b (608 mg, 90%). m NMR (CDCl3) δ 1.24-1.43 (m, 6H), 1.69-2.17 (m, 6H), 2.20-2.38 (m, 6H), 2.48-2.84 (m, 6H), 3.18-3.30 (m, 4H), 3.76-3.83 (m, 4H), 3.95 (s, 3H), 4.09-4.18 (m 2H), 4.62-4.72 (m IH), 4.82-4.84 (d, IH, / = 3.67), 6.78 (s, IH), 7.38-7.46 (t, IH, / = 7.34), 7.64-7-76 (m, 3H), 7.83-7.91 (t, IH, / = 7.34 Hz), 8.69 (s, IH); MS (FAB) 669 [M+H]+.
2S)-Λ/-{4-[4-[4-Piperazinoquinazoline]butyl]oxy]-5-methoxy-2-amino benzoyl} pyrrolidine-2-carboxaldehyde diethyl thioacetal (4b)
The compound 4b has been prepared according to the method described for the compound 4a by employing compound 3b (669 mg, 1 0 mmol) to afford compound 4b (640 mg, 95%), which, due to potential stability problems, was used directly in the next step.
7-Methoxy-8-{4-[4-piperazinoquinazoline]butyl}-oxy-(11aS)-1 ,2,3,11a-5H-pyrro- lo[2,1-c][1 ,4] benzodiazepin-5-one (5b)
The compound 5b was prepared according to the method described for the compound 5a employing the compound 4b (640 mg, 1 0 mmol) to afford the compound 5b (435 mg, 67%).
1H NMR (CDCl3) δ 1.25-1.42 (m, 4H), 1.64-1.88 (m, 4H), 2.02-2.20 (m, 3H), 2.28-2.39 (m, 2H), 2.63-2.78 (m, 4H), 3.80-3.91 (m, 4H), 3.96 (s, 3H), 4.18-4.24 (t, 2H, / = 6.73 Hz), 7.42-7.56 (m, 2H), 7.67-7-79 (m, 2H), 7.87-7.94 (m, 2H), 8.74 (s, IH); MS (FAB) 514 [M+H]+
Example-3
(2S)-A/-{4-[5-[4-piperazinoquinazoline]pentyl]oxy]-5-methoxy-2-nitrobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (3c)
The compound 3c as been prepared according to the method described for the compound 3a by employing the compounds 1c (549 mg, 1 0 mmol) was added anhydrous K2CO3 (690 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3c (629 mg, 92%).
1H NMR (CDCl3) δ 1.22-1.40 (m, 6H), 1.52-1.72 (m, 4H), 1.86-1.99 (m, 6H), 2.00- 2.08 (m, 2H), 2.12-2.18 (m, 4H), 2.21-2.33(m, 2), 3.19-3.30 (m, 4H), 3.32-3.43 (m, 4H), 3.95 (s, 3H), 4.19-4.25 (t, 2H, J = 6.51 Hz), 4.66-4.74 (m, IH), 4.84-4.88 (d, IH, J=3.98 Hz), 6.83 (s, IH), 7.41-7.48 (m, 2H), 7.86-7.93 (m, 3H), 8.73 (s, IH); MS (FAB) 683[M+H]+ (2S)-Λ/-{4-[5-[4-piperazinoquinazoline]pentyl]oxy]-5-methoxy-2-aminobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (4c)
The compound 4c has been prepared according to the method described for the compound 4a by employing compound 3c (683 mg, 1 0 mmol) to afford compound 4c (654 mg, 95%), which, due to potential stability problems, was used directly in the next step.
7-Methoxy-8-{5-[4-piperazinoquinazoline]pentyl}-oxy-(11aS)-1 ,2,3,11a-5/-/-pyrr- olo[2,1-c][1 ,4]benzodiazepin-5-one (5c)
The compound 5c was prepared according to the method described for the compound 5a employing the compound 4c (655 mg, 1 0 mmol) to afford the compound 5c (450 mg, 68%).
1H NMR (CDCl3) δ 1.23-1.44 (m, 4H), 1.62-1.85 (m, 6H), 2.03-2.19 (m, 3H), 2.29-2.38 (m, 2H), 2.62-2.77 (m, 4H), 3.81-3.92 (m, 4H), 3.95 (s, 3H), 4.18-4.25 (t, 2H, / = 6.75 Hz)), 7.44-7.58 (m, 2H), 7.68-7.80 (m, 2H), 7.86-7.93 (m, 2H), 8.75 (s, IH); MS (FAB) 528 [M+H]+
Example-4
(2S)-Λ/-{4-[6-[4-piperazinoquinazoline]hexyl]oxy]-5-methoxy-2-nitrobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (3d)
The compound 3d has been prepared according to the method described for the compound 3a by employing the compounds 1d (563mg, 1 0 mmol) was added anhydrous K2CO3 (690 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3d (655 mg, 89%).
1H NMR (CDCl3) δ 1.23-1.39 (m, 6H), 1.52-1.70 (m, 4H), 1.86-1.98 (m, 4H), 2.00-2.05 (m, 4H), 2.13-2.16 (m, 2H), 2.23-2.32(m, 2H), 2.64-2.85(m, 4H), 3.20-3.30 (m, 4H), 3.32-3.42 (m, 4H), 3.95 (s, 3H), 4.19-4.24 (t, 2H, / = 6.42 Hz), 4.67-4.73 (m, IH), 4.85-4.87 (d, IH, / = 3.96 Hz), 6,82 (s, IH), 7.42-7.47 (m, 2H), 7.86-7.91 (m, 3H), 8.71 (s, IH); MS (FAB) 697 [M+H]+
2S)-Λ/-{4-[6-[4-Piperazinoquinazoline]hexyl]oxy]-5-methoxy-2-aminobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (4d)
The compound 4d has been prepared according to the method described for the compound 4a by employing compound 3d (698 mg, 1 0 mmol) to afford compound 4d (675 mg, 96%), which, due to potential stability problems, was used directly in the next step.
7-Methoxy-8-{5-[4-piperazinoquinazoline]hexyl)-oxy-(11aS)-1 ,2,3,11a-5H-pyrro- lo[2,1-c][1 ,4]benzodiazepin-5-one (5d)
The compound 5d was prepared according to the method described for the compound 5a employing the compound 4d(668 mg, 1 0 mmol) to afford the compound 5d (465 mg, 69%).
1H NMR (CDCl3) δ 1.25-1.42 (m, 6H), 1.64-1.88 (m, 6H), 2.02-2.20 (m, 4H), 2.28-2.39 (m, 2H), 2.63-2.78 (m, 4H), 3.80-3.91 (m, 3H), 3.96 (s, 3H), 4.18-4.24 (t 2H, J = 6.73 Hz)), 7.42-7.56 (m, 2H), 7.67-7-79 (m, 2H), 7.87-7.94 (m, 2H), 8.74 (s, IH); MS (ESI) 542 [M+H]+
Example-5
2S)-Λ/-{4-[8-[4-piperazinoquinazoline]octyl]oxy]-5-methoxy-2-nitrobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (3e)
The compound 3e has been prepared according to the method described for the compound 3a by employing the compounds 1e (563mg, 1 0 mmol) was added anhydrous K2CO3 (712 mg, 5.0 mmol) and 4-piperazino-quinazoline (2a) (214 mg, 1.0 mmol) to afford the compound 3e (675 mg, 90%). m NMR (CDCl3) δ 1.23-1.39 (m, 6H), 1.52-1.70 (m, 6H), 1.86-1.98 (m, 4H), 2.00-2.05 (m, 4H), 2.13-2.16 (m, 2H), 2.23-2.32(m, 2H), 2.64-2.85(m, 4H), 3.20-3.30 (m, 4H), 3.32-3.42 (m, 4H), 3.95 (s, 3H), 4.19-4.24 (t, 2H, / = 6.42 Hz), 4.67-4.73 (m, IH), 4.85-4.87 (d, IH, / = 3.96 Hz), 6,82 (s, IH), 7.42-7.47 (m, 2H), 7.86-7.91 (m, 3H), 8.71 (s, IH); MS (FAB) 697 [M+H]+
2S)-Λ/-{4-[6-[4-Piperazinoquinazoline]octyl]oxy]-5-methoxy-2-aminobenzoyl} pyrrolidine-2-carboxaldehyde diethylthioacetal (4e)
The compound 4e has been prepared according to the method described for the compound 4a by employing compound 3e (714 mg, 1 0 mmol) to afford compound 4e (682 mg, 95%), which, due to potential stability problems, was used directly in the next step.
7-Methoxy-8-{5-[4-piperazinoquinazoline]octyl)-oxy-(11aS)-1 ,2,3,11 a-5/-/-pyrro- lo[2,1-c][1 ,4]benzodiazepin-5-one (5e) The compound 5e was prepared according to the method described for the compound 5a employing the compound 4e(675 mg, 1 0 mmol) to afford the compound 5e (481 mg, 72%).
1H NMR (CDCl3) δ 1.25-1.42 (m, 6H), 1.64-1.88 (m, 8H), 2.02-2.20 (m, 4H)7 2.28-2.39 (m, 2H), 2.63-2.78 (m, 4H), 3.80-3.91 (m, 3H), 3.96 (s, 3H), 4.18-4.24 (t 2H, / = 6.73 Hz)), 7.42-7.56 (m, 2H), 7.67-7-79 (m, 2H), 7.87-7.94 (m, 2H), 8.74 (s, IH); MS (ESI) 556 [M+H]+
Biological Activity: some of in vitro biological activity studies were carried out at the National Cancer Institute, Maryland, USA.
Cytotoxicity:
The compounds (5a) 7-Methoxy-8-{4-[-3-(4-piperazinoquinazoline]propyl}-oxy-
(11 aS)-1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one.
(5b) 7-Methoxy-8-{4-[-4-( 4-piperazinoquinazoline]butyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one.
(5c)7-Methoxy-8-{5-[-3-(4-piperazinoquinazoline]pentyl}-oxy-(11 aS)-1 ,2,3, 11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one.
(5d)7-Methoxy-8-{6-[-3-(4-piperazinoquinazoline]hexyl}-oxy-(11 aS)-1 ,2,3, 11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one.
(5e)7-Methoxy-8-{8-[-3-(4-piperazinoquinazoline]octyl}-oxy-(11aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one were evaluated for in vitro anticancer activity against nine human tumor cells derived from nine cancer types (colon, prostate, oral, lung, cervix and breast cancer) as shown in (Table 1,2 and 3)
5a, 5c and 5d was evaluated for in vitro anticancer activity against sixty human tumor cells derived from nine cancer types ^leukemia, non-small-cell lung, colon, CNS, melanoma, ovarian, prostate, and breast cancer) as shown in (Table 1 and 2). For the compound, dose response curves for each cell line were measured at a minimum of five concentrations at 10 fold dilutions. A protocol of 48 h continuous drug exposure was used and a sulforhodamine B (SRB) protein assay was used to estimate cell viability or growth. The concentration causing 50% cell growth inhibition (GI50), total cell growth inhibition (TGI 0% growth) and 50% cell death (LC50, -50% growth) compared with the control was calculated. The mean graph midpoint values of log10 TGI and log-io LC50 as well as log™ GI50 for 5a is listed in Table 1 and 2). As demonstrated by mean graph pattern, compound 5c and 5d exhibited an interesting profile of activity and selectivity for various cell lines. The mean graph mid point of log-io TGI and log1Q LC50 showed similar pattern to the log-to GI50 mean graph mid points.
Table 1. Log GI50 (concentration in mol/L causing 50% growth inhibition) values for quinazoline-PBD hybrids.
Cancer Cell Lines
5a 5c
5d
Leukemia -5.68 -5.58 -5.49
Non-small cell lung -5.82 -5.38 -4.87
Colon -5.55 -5.76 -5.41
CNS -4.86 -5.61 -4.63
Melanoma -5.65 -5.38 -5.29
Ovarian -5.56 -4.60 -4.28
Renal -5.67 -5.39 -5.17
Prostate -4.63 -5.05 -5.85
Breast -5.57 -4.31 -4.52 each cancer type represents the average of six to eight different cancer cell lines. In vitro evaluation of cytotoxic activity: Among them 5a exhibits a wide spectrum of activity against sixty cell lines in nine cell panels, with GI50 value of <20 nM. In the non-small cell lung cancer panel, the growth of HOP-62, NCI-H23 cell lines were affected by compound 5a with GI50 values as 11.4, 14.6 and 19.7 nM respectively. The GI50 values of compound 5a against colon cancer HCC-2988, HCT-116 and KM12 cell lines are 11.3, 11.8 and 11.7 nM respectively. The Gl50 values for compound 5a against CNS SF-295, SF-539, SNB-19 and SNB-75 cell lines are in a range of 12.6-23.9 nM. Four cancer cell lines (OVCAR-4, OVCAR-5, OVCAR-8 and SK-OV-3) in the ovarian cancer cell panel were affected by compound 5a with Gl50 values of 31.6, 15.1 , 30.5 and 79.8 nM respectively. In this study compound 5c exhibited cytotoxicity activity against renal and breast cancer panels with Gl50 values (11.6-43.4 nM). Compound 5c exhibits activity against sixty cell lines in nine cancer cell panels with GI50 values of <10 μM. Compound 5d exhibits activity against fifty-seven cell lines in nine cancer cell panels, GI50 values of <10 μM. Cytotoxicity of compounds 5a, 5c and 5d in selected cancer cell lines has been illustrated in and 5d have been, illustrated in Table 1, 2 and 3.Table 2. The average GI50 values for each cancer panel of compounds 5a, 5c, and 5d have been illustrated in Table 1, 2 and 3.
Table 2. Cytotoxicit of com ounds 5a, 5c and 5d in selected cancer cell lines
The mean graph mid point values of log™ TGl and log10 LC50 as well as log™ GI50 for 5a, 5c and 5d are listed in Table-3. As demonstrated by mean graph pattern, compounds 5a, 5c and 5d exhibit an interesting profile of activity and selectivity for various cell lines. The mean graph mid points of log™ TGI and log™ LC50 have shown similar pattern to the log™ Gl50 mean graph mid points.
Table-3. log™ Gl50, log™ TGI and log™ LC50 mean graphs midpoints (MG_MID) of In vitro cytotoxicity data for the compounds 5a, 5c and 5d against human tumor cell lines.
Logio Glδo Log™ TGl Log™ LC50
Compound
5a -7.68 -6.66 -4.99
5c -5.68 -4.86 -4.19
5d -6.49 -5.85 -4.96
ADR = Adiramycin is the control drug
Thermal denaturation studies
Compounds were subjected to thermal denaturation studies with duplex-form calf thymus DNA (CT-DNA) using an adaptation of a reported procedure. Working solutions in aqueous buffer (10 mM NaH2PO4ZNa2HPO4, 1 mM Na2EDTA, pH 7.00+0.01) containing CT-DNA (100 μm in phosphate) and the PBD (20 μm) were prepared by addition of concentrated PBD solutions in DMSO to obtain a fixed [PBD]/[DNA] molar ratio of 1 :5. The DNA-PBD solutions were incubated at 37 0C for 0, 18, and 36 h prior to analysis. Samples were monitored at 260 nm using a Beckman DU-7400 spectrophotometer fitted with high performance temperature controller, and heating was applied at 1 0C min"1 in the 40-90 0C range. DNA helix coil transition temperatures (Tm) were obtained from the maxima in the (d>4260)/dr derivative plots. Results are given as the mean ± standard deviation from three determinations and are corrected for the effects of DMSO co-solvent using a linear correction term. Drug-induced alterations in DNA melting behaviour are given by: Δ7m=7m(DNA+PBD)-Tm (DNA alone), where the Tm value for the PBD-free CT- DNA is 69.0 ± 0.01. The fixed [PBD]/[DNA] ratio used did not result in binding saturation of the host DNA duplex for any compound examined. Compound 5a, 5c and 5d at 0 hr, 18 hr and 36 hr gradually increased at 37°C. Table 4. Thermal denaturation data of C8-linked quinazoline hybrids of pyrrolo[2,1-c]- [1 ,4]benzodiazepine with calf thymus (CT) DNA
[PBD]:[DNA] Δrμ (°C)a after incubation at 37 °C for
PBD hybrids molar ratiob O h 18 h
5a 1:5 9.2 10.4
5b 1:5 9.7 10.9
5c 1 :5 10.3 11.8
5d 1 :5 10.6 12.2
1 :5 0.3 0.7
DC-81 a For CT-DNA alone at pH 7.00 ± 0.01 , Tm = 68.5 0C ± 0.01 (mean value from 10 separate determinations), all ΔTm values are ± 0.1-0.20C. For a 1:5 molar ratio of [PBD]/[DNA], where CT-DNA concentration^ 00 μM and ligand concentration = 20 μM in aqueous sodium phosphate buffer [10 mM sodium phosphate + 1 mM EDTA, pH 7.00 ± 0.01]. The Δ7m for PBD hybrids 5a, 5b, 5c and 5d at a [PBD]:[DNA] molar ratio of 1:5 increased to a value of 10.4 0C , 10.9 °C, 11.8 and 12.2 °C after 18 h incubation respectively.
The DNA binding activity for these novel C8-linked quinazoline-PBD hybrids has been examined by thermal denaturation studies using calf thymus (CT) DNA. Melting studies show that these compounds stabilize the thermal helix -> coil or melting stabilization (Δ7" m) for the CT-DNA duplex at pH 7.0, incubated at 37 0C, where PBD/DNA molar ratio is 1 :5. Interestingly, in this assay one of the quinazoline-PBD hybrid (5a-d) elevates the helix melting temperature of CT-DNA by a margin of 12.2 0C after incubation for 18 h at 37 0C. Data for 5a-d and DC-81 are included in Table 4 for comparison.
ADVANTAGES OF THE INVENTION
1. The present invention provides a new pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids useful as antitumour agents.
2. It also provides a process for the preparation of novel pyrrolo[2,1-c][1 ,4] benzodiazepine hybrids

Claims

We claim:
1. Novel quinazoline linked pyrrolo[2,1-c][1, ^benzodiazepine hybrid of general formula 5
Formula 5 wherein, n = 3-6 or 8.and and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O. 2. Novel quinazoline linked pyrrolo [2,1-c][1 ,4] benzodiazepine hybrid according to claim 1 is represented by the group of the following compounds:
7-Methoxy-8-{[-3-(4-pipe'razinoquinazoline]propyl}-oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5a);
7-Methoxy-8-{[-4-( 4-piperazinoquinazoline ]butyl}-oxy-(11aS)-1 ,2,3,11a-5/-/- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5b);
7-Methoxy-8-{[-5-(4-piperazinoquinazoline]pentyl}-oxy-(1 IaS)- 1 ,2,3,11a-5H- pyrrolo[2, 1-c][1 ,4]benzodiazepin-5-one (5c);
7-Methoxy-8-{[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11 aS)~1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5d);
7-Methoxy-8-{[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)-1,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5e);
7-Methoxy-8-{6,7-dimethoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)-
1 ,2,3, 11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5f);
7-Methoxy-8-{6,7-dimethoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)-
1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5g);
7-Methoxy-8-{6,7-dimethoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)-
1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5h);
7-Methoxy-8-{6,7-dimethoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)-
1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5i); 7-Methoxy-8-{6,7-dimethoxy[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)- 1 ,2,3,11a-5H~pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5j; 7-Methoxy-8-{7-ethoxy,6-methoxy[-3-(4-piperazinoquinazoline]propyl}-oxy- (11 aS)-1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5k); 7-Methoxy-8-{7-ethoxy,6-methoxy[-4-(4-piperazinoquinazoline]butyl}-oxy- (11aS)-1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5I); 7-Methoxy-8-{7-ethoxy,6-methoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy- (11aS)-1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5m); 7-Methoxy-8-{7-ethoxy,6-methoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy- (11aS)-1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5n); 7-Methoxy-8-{7-ethoxy,6-methoxy [-8-( 4-piperazinoquinazoline ]octyl}-oxy- (11 aS)-1 ,
2,3, 11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5o).
3. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid according to claim 1 , wherein the structural formula of the representative compound are
4. Quinazoline linked pyrrolo[2,1-c][1,4]benzodiazepine hybrid according to claim 1 , exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung (Hop-62), cervix (SiHa), breast (MCF7, Zr-75-1), colon (Colo205), prostate (DU145, PC3) and oral (DWD, HT1080) cell lines.
5. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against Colo205 for IC50 is in the range of 17 to 80μm, at an exposure period of at least 48 hrs.
6. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids according to claim 1 , wherein the concentration of the compound used for in vitro activity against DU 145 for IC50 is in the range of 16 to 80μm, at an exposure period of at least 48 hrs.
7. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrids according to claim 1 , wherein the concentration of the compound used for in vitro activity against DWD for IC50 is in the range of 6 to 80μm, at an exposure period of at least 48 hrs.
8. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against HoP62 for IC50 is in the range of 13 to 40 μm, at an exposure period of at least 48 hrs.
9. Quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against HT1080 for IC50 is in the range of 6 to 30μm, at an exposure period of at least 48 hrs.
10. Quinazoline linked pyrrolo[2,1-c][1 /^benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against MCF7 for 1C50 is in the range of 27 to about 80μm, at an exposure period of at least 48 hrs.
11. Quinazoline linked pyrrolo[2,1-c][1 /^benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against PC3 for IC50 is in the range of 9 to about 80μm, at an exposure period of at least 48 hrs.
12. Quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against SiHa for IC50 is in the range of 25 to about 80μm, at an exposure period of at least 48 hrs.
13.Quinazoline linked pyrrolo[2,1-c][1 , ^benzodiazepine hybrid according to claim 1 , wherein the concentration of the compound used for in vitro activity against Zr-75-1 for IC50 is in the range of 24 to about 80μm, at an exposure period of at least 48 hrs.
14.A pharmaceutical composition comprising quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid, its derivatives, analogues, salts or mixture thereof optionally with pharmaceutically acceptable carriers, adjuvants and additives.
15.A pharmaceutical composition according to claim 14, wherein the quinazoline linked pyrrolo[2,1-c][1 ,4]benzodiazepine hybrid used is represented by a general formula 5, where n = 3, 4, 5, 6, 8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O
F
Formula 5
16. A process for the preparation of quinazoline linked pyrrolo[2,1-c][1 ^benzodiazepine hybrid of formula 5,
wherein n = 3, 4, 5, 6, 8 and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O the said process comprising the steps of: a) reacting (2S)-Λ/-[4-(n-bromoalkyl)oxy-5-methoxy-2-nitrobenzoyl] pyrrolidine-2-carboxaldehyde diethylthioacetal of formula 1
with substituted and un substituted piperazinoquinazoline derivative selected from the compound of formula 2
and wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=, in the presence of a mild inorganic base , in an aprotic organic solvent, under refluxing temperature to obtain the resultant nitro compound of (2S)-/V-{4-[n~[4- piperazinoquinazoline]aikyl]oxy]-5-methoxy-2-nitrobenzoyl}pyrrolidine-2- carboxaldehyde diethylthioacetal of formula 3a-o.
3a-o wherein R1 and R2 is selected from the group consisting of R1= H, R2=H or R1= CH3O, R2= CH3O or R1= CH3O and R2=C2H5O b) reducing (2S)-Λ/-{4-[n-[4-Piperazinoquinazoline]alkyll]oxy]-5-methoxy-2- nitro benzoyl}pyrrolidine-2-carboxaldehyde diethylthioacetal of formula 3 with SnCI2.2H2O, in an alcohol, under reflux, followed by the evaporation of alcohol and adjusting the pH of the resultant product layer to about 8 by using a base, followed by extraction with ethyl acetate and washing the combined organic phase with brine solution and evaporating the solvent to obtain the desired (2S)-Λ/-{4-[n-[4-
Piperazinoquinazoline]alkyl]oxy]-5-meth oxy-2-amino benzoyl}pyrrolidine- 2-carboxaldehyde diethylthioacetal 4a-o,
4a-o where in R1, FT = H, OCH3, OC2H5
c) reacting (2S)-Λ/-{4-[5-[4-Piperazinoquinazoline]pentyl]oxy]-5-methoxy-2- aminobenzo-yl}pyrrolidine-2-carboxaldehyde diethylthioacetal of formula 4 with mercuric chloride, in a mixture of water and organic solvent, in the presence of mild inorganic basic compound, under stirring, at a temperature of about 20-300C, for a period of 8-12 hrs, followed by the extraction of yellow organic supernatant and washing with sodium bi carbonate and brine, respectively, and evaporating the organic layer, under reduced pressure and further purified by column chromatography to obtain the desired product of quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid of formula 5a-o.
R1 & R2 =H, OCH3 OC2H5
5a-o
17. A process according to claim 16, wherein the mild inorganic base used in steps
(a) is potassium carbonate.
18.A process according to claim 16, wherein the aprotic organic solvent used in step (a) is acetone and acetonitrile.
19. A process according to claim 16, wherein the organic solvent used in step (c) is acetonitrile and acetone.
20. A process according to claim 16, wherein the alcohol used in step (b) is selected from methanol and ethanol.
21. A process according to claim 16, wherein the compounds of formula 5 obtained are represented by a group of the following compounds: 7-Methoxy-8-{[-3-(4-piperazinoquinazoline]propyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5a)
7-Methoxy-8-{[-4-( 4-piperazinoquinazoline ]butyl}-oxy-(11 aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5b);
7-Methoxy-8-{[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)-1 ,2,3,11a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5c);
7-Methoxy-8-{[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)-1 ,2,3,11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5d);
7-Methoxy-8-{[-8-(4-piperazinoquinazoline]octyl}-oxy-(11 aS)-1 ,2,3, 11 a-5H- pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5e);
7-Methoxy-8-{6,7-dimethoxy[-3-(4-piperazinoquinazoline]propyl}-oxy~(11aS)- 1 ,2,3, 11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5f); 7-Methoxy-8-{6,7-dimethoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-(11aS)- 1 ,2,3,11 a-5/-/-pyrrolo[2, 1 -C][1 ,4]benzodiazepin-5-one (5g); 7-Methoxy-8-{6,7-dimethoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-(11aS)- 1 ,2,3,11a-5H-pyrrolo[2,1-c][1,4]benzodiazepin-5-one (5h); 7-Methoxy-8-{6,7-dimethoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-(11aS)-
1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5i);
7-Methoxy-8-{6,7-dimethoxy[-8-(4-piperazinoquinazoline]octyl}-oxy-(11aS)-
1 ,2,3,11a-5H-pyrrolo[2,1-c][1,4]benzodiazepin-5-one (5j);
7-Methoxy-8-{7-ethoxy,6-methoxy[-3-(4-piperazinoquinazoline]propyl}-oxy-
(11 aS)-1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5k);
7-Methoxy-8-{7-ethoxy,6-methoxy[-4-(4-piperazinoquinazoline]butyl}-oxy-
(11aS)-1,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5l);
7-Methoxy-8-{7-ethoxy,6-methoxy[-5-(4-piperazinoquinazoline]pentyl}-oxy-
(11 aS)-1 ,2,3,11 a-5H-pyrrolo[2, 1 -c][1 ,4]benzodiazepin-5-one (5m);
7-Methoxy-8-{7-ethoxy,6-methoxy[-6-(4-piperazinoquinazoline]hexyl}-oxy-
(11 aS)-1 ,2,3, 11 a-5H-pyrrolo[2, 1 -c\[\ ,4]benzodiazepin-5-one (5n);
7-Methoxy-8-{7-ethoxy,6-methoxy [-8-( 4-piperazinoquinazoline ]octyl}-oxy-
(11aS)-1 ,2,3,11a-5H-pyrrolo[2,1-c][1 ,4]benzodiazepin-5-one (5o)
22.A process according to claim 16, wherein quinazoline linked pyrrolo[2,1-c][1 ,4] benzodiazepine hybrid of formula 5a-o exhibits an in vitro anticancer/antitumour activity against human cancer cell lines selected from the group consisting of lung, cervix, breast, colon, prostate and oral cell lines.
EP08873110.4A 2008-03-05 2008-10-31 Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof Not-in-force EP2265613B1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IN518DE2008 2008-03-05
PCT/IN2008/000712 WO2009109984A1 (en) 2008-03-05 2008-10-31 Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof'

Publications (2)

Publication Number Publication Date
EP2265613A1 true EP2265613A1 (en) 2010-12-29
EP2265613B1 EP2265613B1 (en) 2013-07-24

Family

ID=40344785

Family Applications (1)

Application Number Title Priority Date Filing Date
EP08873110.4A Not-in-force EP2265613B1 (en) 2008-03-05 2008-10-31 Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof

Country Status (4)

Country Link
US (1) US8153627B2 (en)
EP (1) EP2265613B1 (en)
JP (1) JP5657395B2 (en)
WO (1) WO2009109984A1 (en)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP6133431B2 (en) 2012-11-24 2017-05-24 ハンジョウ ディーエーシー バイオテック シーオー．，エルティディ．Ｈａｎｇｚｈｏｕ Ｄａｃ Ｂｉｏｔｅｃｈ Ｃｏ．，Ｌｔｄ． Use of hydrophilic conjugates and conjugation reactions between drug molecules and cell binding molecules
US10464955B2 (en) 2014-02-28 2019-11-05 Hangzhou Dac Biotech Co., Ltd. Charged linkers and their uses for conjugation
AU2015242213A1 (en) 2015-07-12 2018-03-08 Hangzhou Dac Biotech Co., Ltd Bridge linkers for conjugation of cell-binding molecules
US9839687B2 (en) 2015-07-15 2017-12-12 Suzhou M-Conj Biotech Co., Ltd. Acetylenedicarboxyl linkers and their uses in specific conjugation of a cell-binding molecule
KR20220147720A (en) 2016-11-14 2022-11-03 항저우 디에이씨 바이오테크 씨오, 엘티디 Conjugation linkers, cell binding molecule-drug conjugates containing the likers, methods of making and uses such conjugates with the linkers

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU2003222444A1 (en) * 2003-03-31 2004-10-25 Council Of Scientific And Industrial Research NEW PYRIMIDINE LINKED PYRROLO(2,1-c)(1,4)BENZODIAZEPINES AS POTENTIAL ANTITUMOUR AGENTS
EP1608663B1 (en) * 2003-03-31 2008-11-05 Council of Scientific and Industrial Research Pyrrolo (2,1-c)(1,4) benzodiazepines dimers as antitumour agents and process thereof
WO2006003670A1 (en) 2004-06-30 2006-01-12 Council Of Scientific And Industrial Research Pyrrolo[2,1-c][1,4]benzodiazepine-napthalimide conjugates linked through piperazine moiety and process for preparation thereof
WO2007054954A1 (en) * 2005-11-10 2007-05-18 Council Of Scientific & Industrial Research A novel pyrrolo[2,1-c][1,4]benzodiazepine hybrid and a process for the preparation thereof
WO2008020456A2 (en) * 2006-08-14 2008-02-21 Council Of Scientific & Industrial Research Pyrrolo[2,1-c][1,4]benzodiazepine hybrids and a process for the preparation thereof

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2009109984A1 *

Also Published As

Publication number Publication date
WO2009109984A1 (en) 2009-09-11
US8153627B2 (en) 2012-04-10
JP5657395B2 (en) 2015-01-21
JP2011513389A (en) 2011-04-28
EP2265613B1 (en) 2013-07-24
US20110046111A1 (en) 2011-02-24

Similar Documents

Publication Publication Date Title
CA2660799C (en) Pyrrolo[2,1-c][1,4]benzodiazepine hybrids and a process for the preparation thereof
US8927538B2 (en) Carbazole linked pyrrolo[2,1-c][1,4]benzodiazefine hybrids as potential anticancer agents and process for the preparation thereof
WO2009109984A1 (en) Quinazoline linked pyrrolo[2,1-c][1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof&#39;
EP2260039B1 (en) Chalcone linked pyrrolo[2,1-c[[1, 4]benzodiazepine hybrids as potential anticancer agents and process for the preparation thereof
US20050222132A1 (en) C8 - linked pyrrolo[2,1-c][1,4]benzodiazepine-acridone/acridine hybrids
EP1608650B1 (en) Pyrene-linked pyrrolo(2,1-c)(1,4)benzodiazepine derivatives useful as anticancer agents
US8722665B2 (en) Cinnamido-pyrrolo[2,1-C][1,4]benzodiazepines as potential anticancer agents and process for the preparation thereof
US20040192677A1 (en) Pyrene-linked pyrrolo[2,1--c][1,4]benzodiazepine hybrids useful as anti-cancer agents
US6683073B1 (en) Pyrimidine linked pyrrolo[2,1-c][1,4]benzodiazepines as potential antitumour agents
EP2271648B1 (en) Isoxazoline linked pyrrolo [2,1-c][1,4]benzodiazepine hybrids as potential anticancer agents and the process for preparattion thereof
EP2350085B1 (en) Quinazolinone linked pyrrolo[2,1 -c][1.4]benzodiazepine hybrids as potential anticancer agents and process forthe preparation thereof
JP2011515459A (en) Benzophenone piperazine-bound pyrrolo [2,1-c] [1,4] benzodiazepine hybrid as anticancer drug candidate and method for preparing the same
EP2648757B1 (en) Pyrrolo[2,1-c][1,4]benzodiazepine linked imidazo[1,5-a]pyridine conjugates as potential antitumour agents and process for the preparation thereof
WO2005063760A1 (en) C-8 LINKED PYRROLO[2,1-c][1,4]BENZODIAZEPINE-ACRIDONE/ACRIDINE HYBRIDS
WO2004087712A1 (en) NEW PYRIMIDINE LINKED PYRROLO[2,1-c][1,4]BENZODIAZEPINES AS POTENTIAL ANTITUMOUR AGENTS

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20100907

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MT NL NO PL PT RO SE SI SK TR

AX Request for extension of the european patent

Extension state: AL BA MK RS

17Q First examination report despatched

Effective date: 20110408

DAX Request for extension of the european patent (deleted)
GRAP Despatch of communication of intention to grant a patent

Free format text: ORIGINAL CODE: EPIDOSNIGR1

GRAP Despatch of communication of intention to grant a patent

Free format text: ORIGINAL CODE: EPIDOSNIGR1

GRAS Grant fee paid

Free format text: ORIGINAL CODE: EPIDOSNIGR3

GRAA (expected) grant

Free format text: ORIGINAL CODE: 0009210

AK Designated contracting states

Kind code of ref document: B1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MT NL NO PL PT RO SE SI SK TR

REG Reference to a national code

Ref country code: GB

Ref legal event code: FG4D

REG Reference to a national code

Ref country code: CH

Ref legal event code: EP

REG Reference to a national code

Ref country code: AT

Ref legal event code: REF

Ref document number: 623375

Country of ref document: AT

Kind code of ref document: T

Effective date: 20130815

REG Reference to a national code

Ref country code: IE

Ref legal event code: FG4D

REG Reference to a national code

Ref country code: DE

Ref legal event code: R096

Ref document number: 602008026316

Country of ref document: DE

Effective date: 20130919

REG Reference to a national code

Ref country code: AT

Ref legal event code: MK05

Ref document number: 623375

Country of ref document: AT

Kind code of ref document: T

Effective date: 20130724

REG Reference to a national code

Ref country code: NL

Ref legal event code: VDEP

Effective date: 20130724

REG Reference to a national code

Ref country code: LT

Ref legal event code: MG4D

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CY

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130918

Ref country code: PT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20131125

Ref country code: LT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: NO

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20131024

Ref country code: BE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: SE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: HR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: AT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: IS

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20131124

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: LV

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: SI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: FI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: GR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20131025

Ref country code: NL

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: PL

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CY

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CZ

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: SK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: EE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: RO

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: DK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: MC

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: ES

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: IT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

PLBE No opposition filed within time limit

Free format text: ORIGINAL CODE: 0009261

REG Reference to a national code

Ref country code: CH

Ref legal event code: PL

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: NO OPPOSITION FILED WITHIN TIME LIMIT

26N No opposition filed

Effective date: 20140425

REG Reference to a national code

Ref country code: IE

Ref legal event code: MM4A

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CH

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20131031

Ref country code: LI

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20131031

REG Reference to a national code

Ref country code: DE

Ref legal event code: R097

Ref document number: 602008026316

Country of ref document: DE

Effective date: 20140425

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: IE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20131031

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: TR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: HU

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT; INVALID AB INITIO

Effective date: 20081031

Ref country code: BG

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

Ref country code: LU

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20131031

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: MT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130724

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 8

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 9

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 10

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 10

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 11

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: GB

Payment date: 20201016

Year of fee payment: 13

Ref country code: FR

Payment date: 20201022

Year of fee payment: 13

Ref country code: DE

Payment date: 20201022

Year of fee payment: 13

REG Reference to a national code

Ref country code: DE

Ref legal event code: R119

Ref document number: 602008026316

Country of ref document: DE

GBPC Gb: european patent ceased through non-payment of renewal fee

Effective date: 20211031

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: GB

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20211031

Ref country code: DE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20220503

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: FR

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20211031